PROJECT SUMMARY Sleep is a fundamental biological process that is essential for survival. Sleep abnormalities not only affect daily performance but also contribute to various diseases, including cognitive disorders and cardiovascular diseases. It is estimated that 50-70 million Americans suffer from chronic sleep loss and other sleep disorders, such as insomnia. However, effective treatments for sleep disorders are limited. Thus, it is imperative to understand the genetic basis, neural circuits and functions of sleep. Recent studies suggest that sleep is an evolutionarily conserved process, with shared features across different organisms that include behavioral quiescence, increased arousal threshold, and rapid reversibility to wakefulness. To understand the conserved mechanisms underlying sleep regulation and function, I will study a robust sleep model (EGF-induced sleep in C. elegans) that we have generated. It has been shown that epidermal growth factor (EGF) signaling promotes sleep in worms, flies, fish and mammals. In C. elegans, transient activation of EGF signaling in the neuroendocrine cell (ALA) promotes a sleep-like state. My preliminary data shows that transcriptional changes occur during EGF-induced sleep. However, the identities of sleep-promoting neurons downstream of ALA and the molecular mechanisms underlying the impacts of EGF-induced sleep on the worm?s physiology are still unknown. I hypothesize that ALA coordinates downstream sleep control center(s) to drive a sleep state, which induces transcriptional changes to impact physiology. To facilitate the identification of these downstream sleep-promoting neurons, I have developed a new bipartite expression system, called cGAL, for spatiotemporal control of transgene expression in C. elegans, similar to the GAL4-UAS system for Drosophila. With this new genetic tool, I propose to: 1) Identify the neural circuits underlying EGF-induced sleep; 2) Refine cGAL for better spatial and temporal control of transgene expression in C. elegans; 3) Determine transcriptional changes during EGF-induced sleep. To achieve these aims, I need additional scientific and technical training on RNA-seq, bioinformatics, optogenetics, microscopy and circuit mapping the bipartite cGAL system. The K99/R00 award will allow me to acquire these skills with the guidance from my mentors (Dr. Paul Sternberg and Dr. David Prober) and my advisory committee (Dr. Barbara Wold, Dr. David Anderson, Dr. Viviana Gradinanu, Dr. Andres Collazo and Dr. Igor Antoshechkin).The results of this proposal will provide a framework for my future career to focus on the study of the molecular and cellular mechanisms underlying sleep regulation and function in my own laboratory. My study will advance our understanding of the biology of sleep, which may ultimately contribute to the development of effective treatments for sleep disorders. In addition, this project will also provide the C. elegans community with a fully functional bipartite expression system through a valuable set of cGAL reagents, which will allow other researchers to dissect the neural circuits for other complex behaviors in C. elegans.